Control of mRNA translation is an important mechanism for the temporal and spatial regulation of gene expression that underlies the development of an organism. Translational regulation of mRNAs involved in a variety of developmental processes, including oocyte polarization, cell cycle regulation, embryonic patterning and cell fate determination, has been shown to depend on sequences found within their 3' untranslated regions (3'UTRs). The mechanisms by which these 3'UTR regulatory elements and their cognate binding factors control the translational machinery is poorly understood, however. Translational repression of nanos RNA plays an essential role in generating the restricted distribution of Nanos protein that is necessary for proper patterning of the anterior-posterior body axis of the Drosophila embryo. Repression of nanos RNA is mediated by two stem-loops that contribute to a 90 nucleotide translational control element (TCE) within the nanos 3'UTR. The proposed work focuses on translational repression of nanos RNA in Drosophila as a model for investigation of 3'UTR-dependent translational regulatory mechanisms using biochemical, molecular, and genetic approaches. These studies will lead to an understanding of how 3'UTR sequences regulate translation during development. More generally, they will shed light on mechanisms by which RNA-protein interactions provide the highly selective control of basic cellular processes needed for development, growth, and differentiation. Specific Aim 1 combines biochemical and genetic experiments designed to determine how the two TCE stem-loops and their binding factors block translation of nanos RNA. Aim 2 focuses on biochemical and genetic characterization of a recently identified TCE-binding factor and isolation of interacting proteins, to determine its function in nanos regulation. Aim 3 provides a complementary approach, using a directed genetic screen to isolation of genes encoding components of the regulatory machinery, including those that do not interact directly with the TCE as well as those that have pleiotropic roles during development.